Activation Of GABAA Receptors Research Articles

Taurine Promotes Differentiation and Maturation of Neural Stem/Progenitor Cells from the Subventricular Zone via Activation of GABAA Receptors

Neurogenesis, the formation of new neurons in the brain, occurs throughout the lifespan in the subgranular zone of the dentate gyrus and subventricular zone (SVZ) lining the lateral ventricles of the mammal brain. In this process, gamma-aminobutyric acid (GABA) and its ionotropic receptor, the GABAA receptor (GABAAR), play a critical role in the proliferation, differentiation, and migration process of neural stem/progenitor cells (NPC). Taurine, a non-essential amino acid widely distributed throughout the central nervous system, increases the proliferation of SVZ progenitor cells by a mechanism that may involve GABAAR activation. Therefore, we characterized the effects of taurine on the differentiation process of NPC expressing GABAAR. Preincubation of NPC-SVZ with taurine increased microtubule-stabilizing proteins assessed with the doublecortin assay. Taurine, like GABA, stimulated a neuronal-like morphology of NPC-SVZ and increased the number and length of primary, secondary, and tertiary neurites compared with control NPC of the SVZ. Furthermore, neurite outgrowth was prevented when simultaneously incubating cells with taurine or GABA and the GABAAR blocker, picrotoxin. Patch-clamp recordings revealed a series of modifications in the NPCs’ passive and active electrophysiological properties exposed to taurine, including regenerative spikes with kinetic properties similar to the action potentials of functional neurons.

Acute deep brain stimulation of the paraventricular nucleus of the hypothalamus increases brown adipose tissue thermogenesis in rats

Brown adipose tissue (BAT) activity is controlled by the sympathetic nervous system. Activation of BAT has shown significant promise in preclinical studies to elicit weight loss. Since the hypothalamic paraventricular nucleus (PVN) contributes to the regulation of BAT thermogenic activity, we sought to determine the effects of electrical stimulation of the PVN as a model of deep brain stimulation (DBS) for increasing BAT sympathetic nerve activity (SNA). The rostral raphe pallidus area (rRPa) was also chosen as a target for DBS since it contains the sympathetic premotor neurons for BAT. Electrical stimulation (100 µA, 100 µs, 100 Hz, for 5 min at a 50 % duty cycle) of the PVN increased BAT SNA and BAT thermogenesis. These effects were prevented by a local nanoinjection of bicuculline, a GABAA receptor antagonist. We suggest that electrical stimulation of the PVN elicited local release of GABA, which inhibited BAT sympathoinhibitory neurons in PVN, thereby releasing a restraint on BAT SNA. Electrical stimulation of the rRPa inhibited BAT thermogenesis and this was prevented by a local nanoinjection of bicuculline, suggesting that local release of GABA suppressed BAT SNA. Electrical stimulation of the PVN activates BAT metabolism via a mechanism that may include activation of local GABAA receptors. These findings contribute to our understanding of the mechanisms underlying the effects of DBS in the regulation of fat metabolism and provide a foundation for further DBS studies targeting hypothalamic circuits regulating BAT thermogenesis as a therapy for obesity.

The GABA and GABA-Receptor System in Inflammation, Anti-Tumor Immune Responses, and COVID-19

GABA and GABAA-receptors (GABAA-Rs) play major roles in neurodevelopment and neurotransmission in the central nervous system (CNS). There has been a growing appreciation that GABAA-Rs are also present on most immune cells. Studies in the fields of autoimmune disease, cancer, parasitology, and virology have observed that GABA-R ligands have anti-inflammatory actions on T cells and antigen-presenting cells (APCs), while also enhancing regulatory T cell (Treg) responses and shifting APCs toward anti-inflammatory phenotypes. These actions have enabled GABAA-R ligands to ameliorate autoimmune diseases, such as type 1 diabetes (T1D), multiple sclerosis (MS), and rheumatoid arthritis, as well as type 2 diabetes (T2D)-associated inflammation in preclinical models. Conversely, antagonism of GABAA-R activity promotes the pro-inflammatory responses of T cells and APCs, enhancing anti-tumor responses and reducing tumor burden in models of solid tumors. Lung epithelial cells also express GABA-Rs, whose activation helps maintain fluid homeostasis and promote recovery from injury. The ability of GABAA-R agonists to limit both excessive immune responses and lung epithelial cell injury may underlie recent findings that GABAA-R agonists reduce the severity of disease in mice infected with highly lethal coronaviruses (SARS-CoV-2 and MHV-1). These observations suggest that GABAA-R agonists may provide off-the-shelf therapies for COVID-19 caused by new SARS-CoV-2 variants, as well as novel beta-coronaviruses, which evade vaccine-induced immune responses and antiviral medications. We review these findings and further advance the notions that (1) immune cells possess GABAA-Rs to limit inflammation in the CNS, and (2) this natural "braking system" on inflammatory responses may be pharmacologically engaged to slow the progression of autoimmune diseases, reduce the severity of COVID-19, and perhaps limit neuroinflammation associated with long COVID.

HDAC1-mediated regulation of GABA signaling within the lateral septum facilitates long-lasting social fear extinction in male mice

Social anxiety disorder (SAD) is caused by traumatic social experiences. It is characterized by intense fear and avoidance of social contexts, which can be robustly mimicked by the social fear conditioning (SFC) paradigm. The extinction phase of the SFC paradigm is akin to exposure therapy for SAD and requires learning to disassociate the trauma with the social context. Learning-induced acetylation of histones is critical for extinction memory formation and its endurance. Although class I histone deacetylases (HDACs) regulate the abovementioned learning process, there is a lack of clarity in isoforms and spatial specificity in HDAC function in social learning. Utilizing the SFC paradigm, we functionally characterized the role of HDAC1, specifically in the lateral septum (LS), in regulating the formation of long-term social fear extinction memory. We measured a local increase in activity-inducing HDAC1 phosphorylation at serine residues of social fear-conditioned (SFC+) mice in response to the extinction of social fear. We also found that LS-HDAC1 function negatively correlates with acute social fear extinction learning using pharmacological and viral approaches. Further, inhibition of LS-HDAC1 enhanced the expression of the GABA-A receptor β1 subunit (Gabrb1) in SFC+ mice, and activation of GABA-A receptors facilitated acute extinction learning. Finally, the facilitation of extinction learning by HDAC1 inhibition or GABA-A receptor activation within the LS led to the formation of long-lasting extinction memory, which persisted even 30 days after extinction. Our results show that HDAC1-mediated regulation of GABA signaling in the LS is crucial for the formation of long-lasting social fear extinction memory.

The delta subunit of the GABAA receptor is necessary for the GPT2-promoted breast cancer metastasis.

Objectives: Glutamic pyruvate transaminase (GPT2) catalyzes the reversible transamination between alanine and α-ketoglutarate (α-KG) to generate pyruvate and glutamate during cellular glutamine catabolism. The glutamate could be further converted to γ-aminobutyric acid (GABA). However, the role of GPT2 in tumor metastasis remains unclear. Methods: The wound healing and transwell assays were carried out to analyze breast cancer cell migration and invasion in vitro. Gene ontology analysis was utilized following RNA-sequencing to discover the associated molecule function. The mass spectrometry analysis following phosphoprotein enrichment was performed to discover the associated transcription factors. Most importantly, both the tail vein model and Mammary gland conditional Gpt2-/- spontaneous tumor mouse models were used to evaluate the effect of GPT2 on breast cancer metastasis in vivo. Results: GPT2 overexpression increases the content of GABA and promotes breast cancer metastasis by activating GABAA receptors. The delta subunit GABRD is necessary for the GPT2/GABA-induced breast cancer metastasis in xenograft and transgenic mouse models. Gpt2 knockout reduces the lung metastasis of the genetic Gpt2-/- breast cancer in mice and prolongs the overall survival of tumor burden mice. Mechanistically, GPT2-induced GABAA receptor activation increases Ca2+ influx by turning on its associated calcium channel, and the surged intracellular calcium triggers the PKC-CREB pathway activation. The activated transcription factor CREB accelerates breast cancer metastasis by upregulating metastasis-related gene expressions, such as PODXL, MMP3, and MMP9. Conclusion: In summary, this study demonstrates that GPT2 promotes breast cancer metastasis through up-regulated GABA activation of GABAAR-PKC-CREB signaling, suggesting it is a potential target for breast cancer therapy.

Hybrid molecules combining GABA-A and serotonin 5-HT6 receptors activity designed to tackle neuroinflammation associated with depression

There is clear evidence that the presence of inflammatory factors and impaired GABA-ergic neurotransmission in depressed patients is associated with poor clinical outcome. We designed hybrid molecules, bearing the GABA molecule assembled with chemical fragments that interact with the serotonin 5-HT6 receptor. Such a combination aimed to curb neuroinflammation, remodel GABA-ergic signaling, and provide antidepressant-like activity. The most promising hybrid 3B exerted nanomolar affinity for 5-HT6 receptors and exerted agonistic properties on GABA-A receptors. Developability studies conferred that 3B exerted favorable drug-like properties and optimal brain penetration. In in vivo studies, 3B exerted robust antidepressant-like activity and proved to be highly effective in reducing levels of oxidative stress markers and the pro-inflammatory cytokine IL-6. The inetersting pharmacological profile of 3B makes it a promising candidate for further development for depression associated with neuroinflammation.

14-3-3ζ Mediates GABAAR Activation by Interacting with BIG1.

Most fast synaptic inhibitions in the mammalian brain are mediated by GABAA receptors (GABAARs). An appropriate level of GABAAR expression at the cell surface is essential for neurodevelopment and the efficacy of GABAergic synaptic transmission. We previously reported that brefeldin A-inhibited GDP/GTP exchange factor 1 (BIG1), a binding partner of GABAARs, plays an important role in trafficking GABAARs to the cell surface. However, its regulatory mechanisms remain unknown. In the present study, we identified a new cellular protein, 14-3-3ζ, which can interact with the β subunit of GABAARs and BIG1 both in vitro and in vivo and colocalizes in the soma, dendrites, and axons of hippocampal neurons. Overexpression of 14-3-3ζ-WT increased the surface expression of BIG1 in dendrites and axons, as well as the binding of BIG1 with GABAAR. Depleted 14-3-3ζ with efficacious siRNA attenuated the interaction between BIG1 and GABAARs and resulted in significant decreases in the surface expression levels of BIG1 and GABAAR. GABAAR agonist treatment increased the expression levels of BIG1 and 14-3-3ζ on the surface, indicating that 14-3-3ζ is involved in regulating BIG1-mediated GABAAR surface expression. Depletion of BIG1 or 14-3-3ζ significantly decreased GABAAR expression at the cell surface and suppressed the GABA-gated influx of chloride ions. These data indicate that the combination of 14-3-3ζ and BIG1 is required for GABAAR membrane expression. Our results provide a potential promising therapeutic target for neurological disorders involving GABAergic synaptic transmission.

Dexmedetomidine Attenuates Lipopolysaccharide-Induced Sympathetic Activation and Sepsis via Suppressing Superoxide Signaling in Paraventricular Nucleus.

Sympathetic overactivity contributes to the pathogenesis of sepsis. The selective α2-adrenergic receptor agonist dexmedetomidine (DEX) is widely used for perioperative sedation and analgesia. We aimed to determine the central roles and mechanisms of DEX in attenuating sympathetic activity and inflammation in sepsis. Sepsis was induced by a single intraperitoneal injection of lipopolysaccharide (LPS) in rats. Effects of DEX were investigated 24 h after injection of LPS. Bilateral microinjection of DEX in the paraventricular nucleus (PVN) attenuated LPS-induced sympathetic overactivity, which was attenuated by the superoxide dismutase inhibitor DETC, cAMP analog db-cAMP or GABAA receptor antagonist gabazine. Superoxide scavenger tempol, NADPH oxidase inhibitor apocynin, adenylate cyclase inhibitor SQ22536 or PKA inhibitor Rp-cAMP caused similar effects to DEX in attenuating LPS-induced sympathetic activation. DEX inhibited LPS-induced superoxide and cAMP production, as well as NADPH oxidase, adenylate cyclase and PKA activation. The roles of DEX in reducing superoxide production and NADPH oxidase activation were attenuated by db-cAMP or gabazine. Intravenous infusion of DEX inhibited LPS-induced sympathetic overactivity, NOX activation, superoxide production, TNF-α and IL-1β upregulation in the PVN and plasma, as well as lung and renal injury, which were attenuated by the PVN microinjection of yohimbine and DETC. We conclude that activation of α2-adrenergic receptors with DEX in the PVN attenuated LPS-induced sympathetic overactivity by reducing NADPH oxidase-dependent superoxide production via both inhibiting adenylate cyclase-cAMP-PKA signaling and activating GABAA receptors. The inhibition of NADPH oxidase-dependent superoxide production in the PVN partially contributes to the roles of intravenous infusion of DEX in attenuating LPS-induced sympathetic activation, oxidative stress and inflammation.

Mifepristone blocks the anxiolytic- and antidepressant-like effects of allopregnanolone in male rats

Background Allopregnanolone (3α, 5α-tetrahydroprogesterone) is an inhibitory neurosteroid synthesized from progesterone via 5α-reductase activity in the brain and has anxiolytic, antidepressant, sedative, anticonvulsant, and analgesic activity. Altered levels of allopregnanolone cause anxiety, depression, premenstrual syndrome, and psychiatric disorders. Although allopregnanolone exerts most of its actions by modulating GABAA receptor, NMDA receptor, BDNF expression, and PXR activity, a recent study showed its effects are blocked by mifepristone on lordosis behavior which indicates the involvement of progestin or glucocorticoid receptors in the effects of allopregnanolone since mifepristone blocks both these receptors. However, whether these receptors are involved in acute anxiolytic or antidepressant-like effects is unknown. Methods Adult male Wistar rats were used to study whether the prior administration of mifepristone would alter the effects of allopregnanolone in the elevated plus maze (EPM) and forced swim test (FST) was evaluated. Results 10 mg/Kg dose of allopregnanolone increased percent open arm entries in the EPM, whereas 3 mg/Kg dose of allopregnanolone decreased percent immobility in the FST. Mifepristone administration resulted in a U-shaped response in the FST (with 1 mg/Kg, s.c., decreasing the immobility time) without significantly impacting the behavior in the EPM. In combination studies, mifepristone blocked the anxiolytic and antidepressant effects of allopregnanolone. Conclusion The current study provides evidence for the first time that progestin or glucocorticoid receptors are involved in the acute anxiolytic and antidepressant effects of allopregnanolone. Understanding the mechanism of action of allopregnanolone will help us design better therapeutic strategies to treat neuropsychiatric diseases such as depression and anxiety.

Neuroprotective effect of Ziziphi Spinosae Semen on rats with p-chlorophenylalanine-induced insomnia via activation of GABAA receptor.

Ziziphus jujuba var. spinosa (Bunge) Hu ex H.F.Chow [Rhamnaceae; Ziziphi Spinosae Semen (ZSS)] has attracted extensive attention as the first choice of traditional Chinese medicine in the treatment of insomnia. However, recent studies on the sleep-improving mechanism of ZSS have mainly focused on the role of single components. Thus, to further reveal the potential mechanism of ZSS, an assessment of its multiple constituents is necessary. In this study, ZSS extract (ZSSE) was obtained from ZSS via detailed modern extraction, separation, and purification technologies. The chemical constituents of ZSSE were analyzed by high-performance liquid chromatography-mass spectrometry (HPLC-MS). For in vivo experiments, a rat model of insomnia induced by p-chlorophenylalanine (PCPA) was established to investigate the potential effect and corresponding mechanism of ZSSE on improving sleep. Hematoxylin-eosin staining (HE) results revealed that the drug group showed prominent advantages over the model group in improving sleep. Moreover, the brain levels of γ-aminobutyric acid (GABA), glutamic acid (Glu), 5-hydroxytryptamine (5-HT), and dopamine (DA) were monitored via enzyme-linked immunosorbent assay (ELISA) to further study the sleep-improving mechanism of ZSSE. We found that sleep was effectively improved via upregulation of GABA and 5-HT and downregulation of Glu and DA. In addition, molecular mechanisms of ZSSE in improving sleep were studied by immunohistochemical analysis. The results showed that sleep was improved by regulating the expression levels of GABA receptor subunit alpha-1 (GABAARα1) and GABA acid receptor subunit gamma-2 (GABAARγ2) receptors in the hypothalamus and hippocampus tissue sections. Therefore, this work not only identified the active ingredients of ZSSE but also revealed the potential pharmacological mechanism of ZSSE for improving sleep, which may greatly stimulate the prospective development and application of ZSSE.

Sex-specific hypnotic effects of the neuroactive steroid (3β,5β,17β)-3-hydroxyandrostane-17-carbonitrile are mediated by peripheral metabolism into an active hypnotic steroid

BackgroundThe novel synthetic neuroactive steroid (3β,5β,17β)-3-hydroxyandrostane-17-carbonitrile (3β-OH) blocks T-type calcium channels but does not directly modulate neuronal γ-aminobutyric acid type A (GABAA) currents like other anaesthetic neurosteroids. As 3β-OH has sex-specific hypnotic effects in adult rats, we studied the mechanism contributing to sex differences in its effects. MethodsWe used a combination of behavioural loss of righting reflex, neuroendocrine, pharmacokinetic, in vitro patch-clamp electrophysiology, and in vivo electrophysiological approaches in wild-type mice and in genetic knockouts of the CaV3.1 T-type calcium channel isoform to study the mechanisms by which 3β-OH and its metabolite produces sex-specific hypnotic effects. ResultsAdult male mice were less sensitive to the hypnotic effects of 3β-OH compared with female mice, and these differences appeared during development. Adult males had higher 3β-OH brain concentrations despite being less sensitive to its hypnotic effects. Females metabolised 3β-OH into the active GABAA receptor positive allosteric modulator (3α,5β,17β)-3-hydroxyandrostane-17-carbonitrile (3α-OH) to a greater extent than males. The 3α-OH metabolite has T-channel blocking properties with sex-specific hypnotic and pharmacokinetic effects. Sex-dependent suppression of the cortical electroencephalogram is more pronounced with 3α-OH compared with 3β-OH. ConclusionsThe sex-specific differences in the hypnotic effect of 3β-OH in mice are attributable to differences in its peripheral metabolism into the more potent hypnotic metabolite 3α-OH.

Sedative-hypnotic effects of Boropinol-B on mice via activation of GABAA receptors.

Boropinol-B is a phenylpropanoid compound originally isolated from Boronia pinnata Sm. (Rutaceae). This study aimed to evaluate the sedative-hypnotic effects of Boropinol-B and explore the underlying mechanisms. Pentobarbital sodium-induced sleep mouse model and caffeine-induced insomnia mouse model were used to investigate the sedative effects of Boropinol-B. Pharmacokinetics profiles of Boropinol-B in rats were evaluated by high-performance liquid chromatography. The effects of Boropinol-B on the γ-aminobutyric acid (GABA)ergic system were investigated using ELISA assay and patch-clamp technique. Immunohistochemistry and immunofluorescence were carried out to assess the effects of Boropinol-B on sleep-related brain nucleus. Boropinol-B showed significant sedative effects, including reduced sleep latency, increased sleep duration in pentobarbital sodium-treated mice and decreased locomotor activity in insomnia mice. Pharmacokinetics studies demonstrated that Boropinol-B had a rapid onset of action, a short half-life and no accumulation. It increased the GABA level in mice's brain, and promoted chloride ions influx mediated by the γ-aminobutyric acid type A (GABAA) receptors in neurons. Also, it increased the c-Fos positive ratio of GABAergic neurons in ventrolateral preoptic nucleus and decreased c-Fos expression in tuberomammillary nucleus. Boropinol-B showed significant sedative-hypnotic effects in mice by activating the GABAA receptors and stimulating the sleep-related brain nucleus.

Possible Interaction of Opioidergic and Nitrergic Pathways in the Anticonvulsant Effect of Ivermectin on Pentylenetetrazole-Induced Clonic Seizures in Mice.

Ivermectin (IVM) is an antiparasitic drug that primarily works by the activation of GABAA receptors. The potential pharmacological pathways behind the anti-convulsant effect of IVM haven't yet been identified. In this study, intravenous injection of pentylenetetrazole (PTZ)-induced clonic seizure in mice was investigated in order to assess the possible influence of IVM on clonic seizure threshold (CST). We also look at the function of the Opioidergic and nitrergic pathways in IVM anticonvulsant action on clonic seizure threshold. IVM (0.5, 1, 5, and 10mg/kg, i.p.) raised the PTZ-induced CST, according to our findings. Furthermore, the ineffective dose of nitric oxide synthase inhibitors (L-NAME 10mg/kg, i.p.), and (7-NI 30mg/kg, i.p.) or opioidergic system agonist (morphine 0.25mg/kg, i.p.) were able to amplify the anticonvulsive action of IVM (0.2mg/kg, i.p.). Moreover, the anticonvulsant effect of IVM was reversed by an opioid receptor antagonist (naltrexone 1mg/kg, i.p.). Furthermore, the combination of the ineffective dose of morphine as an opioid receptor agonist with either L-NAME (2mg/kg, i.p.) or 7-NI (10mg/kg, i.p.) and with an ineffective dose of IVM (0.2mg/kg, i.p.) had a significant anticonvulsant effect. Taken together, IVM has anticonvulsant activity against PTZ-induced clonic seizures in mice, which may be mediated at least in part through the interaction of the opioidergic system and the nitric oxide pathway.

RBIO-02. ACTIVATION OF GABAA RECEPTORS WITH A NON-TOXIC, BRAIN PENETRANT SMALL MOLECULE SENSITIZES LUNG ADENOCARCINOMA PRIMARY AND BRAIN METASTATIC TUMOR CELLS TO RADIATION VIA AUTOPHAGY INDUCTION

Abstract Most advanced-stage non-small cell lung cancer (NSCLC) patients have brain metastases that render a dismal prognosis. Treatment of metastatic brain lesions from NSCLC and other tumor types include radiation as part of a multimodal treatment regimen. Challenges in the application of radiotherapy include overcoming radiation resistance and reducing associated co-morbidities. Non-toxic therapeutics capable of sensitizing tumors to radiation are needed to improve survival and mitigate radiation side-effects. Many CNS and solid systemic tumors express ligand-gated ion channels, which may contribute to tumor growth. Leveraging ion channels is therefore a potential way of diminishing the spread of cancer. We find that NSCLC and its brain metastases express subunits of the type-A GABA-gated chloride channel or GABAA receptor. Importantly, patient-derived NSCLC cells have functional GABAA receptors. We identified a brain penetrant, small molecule activator of GABAA receptors (AMLAL-101), which alone impairs the viability of both primary NSCLC cells and brain metastatic cells. In addition, AMLAL-101 combined with radiation is a highly potent inducer of NSCLC cell death and clonogenic arrest. Using a human ex vivo model of NSCLC-on-chip, we assessed the efficacy and toxicity of AMLAL-101 relative to Docetaxel, an antimicrotubular agent used in treating advanced NSCLC. AMLAL-101 is as potent as Docetaxel but does not exhibit its toxic side effects. AMLAL-101 also potentiates radiation in vivo, significantly reducing lung adenocarcinoma xenograft tumor growth in mice, equivalent to docetaxel plus radiation. Mechanistically, AMLAL-101 activates GABAA receptors in NSCLC and synergizes with radiation by inducing an autophagic response that includes: (i) stabilization of Beclin-1, BNIP3L/NIX, and GABARAP; (ii) ATG7 upregulation; and (iii) utilization of ubiquitin-binding protein p62. Activating GABAA receptors in NSCLC and other tumor types may improve radiation efficacy and mitigate its toxic side effects in treating brain metastases.

RBIO-01. ACTIVATING AN ION CHANNEL PROMOTES RADIATION-SENSITIZING IMMUNE-DRIVEN ANTI-TUMOR ACTIVITY

Abstract Standard-of-care for melanoma brain metastases is moving to stereotactic radiosurgery (SRS) with immunotherapy. Although this combined approach shows improved patient survival, the responses are not durable. SRS can also cause adverse reactions such as radiation necrosis, which can result in significant morbidity and even mortality. Lowering SRS dose without negatively impacting its effectiveness would therefore be highly beneficial clinically. Our published data suggests that when treating syngeneic mouse model B16F10 melanoma tumors, the radiation dose can be reduced and its effectiveness can be improved, by activating intrinsic melanoma GABAA receptor (GABAAR) activity using a novel agent, AMLAL-101. This agent acts to enhance GABAAR mediated chloride transport in melanoma cells, which in turn triggers mitochondrial dysregulation associated with tumor regression in mice. While AMLAL-101 alone reduces tumor growth, combining it with radiation results in near complete regression of the irradiated tumor, as well as shrinking of tumor not in the radiation field due to an immune response, i.e., a potent abscopal effect. AMLAL-101 may also function to regulate immune cell infiltration into the tumor microenvironment. Analysis of AMLAL-101 treated tumors show increased infiltration into the tumor microenvironment of: (1) polyfunctional (TNFα/IFN-γ) CD8+ T cells; and (2) tumor-associated macrophages (TAMs) of the tumor inhibitory M1-phenotype, possibly due to IFN-γ stimulation. We posit that activating GABAAR with AMLAL-101 promotes both a ‘direct’ anti-tumor response and an ‘indirect’, immune-driven anti-tumor activity in metastatic melanoma tumors. AMLAL-101 may address an urgent unmet need for a non-toxic therapeutic that can potentiate radiation and mitigate its toxicity profile to positively impact clinical outcomes of patients with brain metastatic disease treated with radiotherapy.

Pharmacological activation of mediodorsal thalamic GABA-A receptors modulates morphine/cetirizine-induced changes in the prefrontal cortical GFAP expression in a rat model of neuropathic pain

The present study investigated the involvement of mediodorsal thalamic (MD) GABA-A receptors in cetirizine/morphine-induced anti-allodynia using a rat model of neuropathic pain. To assess the importance of the prefrontal cortex (PFC) for chronic pain processing, its expression level changes of glial fibrillary acidic protein (GFAP) were measured following drug treatments. Each animal was subjected to chronic constriction of the sciatic nerve surgery simultaneously with the MD cannulation under stereotaxic surgery. The results showed that the administration of morphine (3–5 mg/kg) or cetirizine (1–3 mg/kg) produced significant analgesia in neuropathic rats. Systemic administration of cetirizine (2.5 and 3 mg/kg) potentiated the analgesic response to a low and intolerance dose of morphine (3 mg/kg). Intra-MD microinjection of muscimol, a selective GABA-A receptor agonist (0.005–0.01 μg/rat), increased the cetirizine/morphine-induced anti-allodynia, while muscimol by itself did not affect neuropathic pain. The neuropathic pain was associated with the increased PFC expression level of GFAP, suggesting the impact of chronic pain on PFC glial management. Interestingly, the anti-allodynia was associated with a decrease in the PFC expression level of GFAP under the drugs' co-administration. Thus, cetirizine has a significant potentiating effect on morphine response in neuropathic pain via interacting with the MD GABA-A receptors. It seems that neuropathic pain affects the prefrontal cortex GFAP signaling pathway. In clinical studies, these findings can be considered to create a combination therapy with low doses of GABA-A receptor agonist plus cetirizine and morphine to manage neuropathic pain.

GABAergic Regulation of Astroglial Gliotransmission through Cx43 Hemichannels

Gamma-Aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the brain. It is produced by interneurons and recycled by astrocytes. In neurons, GABA activates the influx of Cl- via the GABAA receptor or efflux or K+ via the GABAB receptor, inducing hyperpolarization and synaptic inhibition. In astrocytes, the activation of both GABAA and GABAB receptors induces an increase in intracellular Ca2+ and the release of glutamate and ATP. Connexin 43 (Cx43) hemichannels are among the main Ca2+-dependent cellular mechanisms for the astroglial release of glutamate and ATP. However, no study has evaluated the effect of GABA on astroglial Cx43 hemichannel activity and Cx43 hemichannel-mediated gliotransmission. Here we assessed the effects of GABA on Cx43 hemichannel activity in DI NCT1 rat astrocytes and hippocampal brain slices. We found that GABA induces a Ca2+-dependent increase in Cx43 hemichannel activity in astrocytes mediated by the GABAA receptor, as it was blunted by the GABAA receptor antagonist bicuculline but unaffected by GABAB receptor antagonist CGP55845. Moreover, GABA induced the Cx43 hemichannel-dependent release of glutamate and ATP, which was also prevented by bicuculline, but unaffected by CGP. Gliotransmission in response to GABA was also unaffected by pannexin 1 channel blockade. These results are discussed in terms of the possible role of astroglial Cx43 hemichannel-mediated glutamate and ATP release in regulating the excitatory/inhibitory balance in the brain and their possible contribution to psychiatric disorders.

Neuroprotective and anti-epileptic potentials of genus Artemisia L.

The Genus Artemisia L. is one of the largest genera in the Asteraceae family growing wild over in Europe, North America, and Central Asia and has been widely used in folk medicine for the treatment of various ailments. Phytochemical and psychopharmacological studies indicated that the genus Artemisia extracts contain various antioxidant and anti-inflammatory compounds and possess antioxidant, anti-inflammatory, antimicrobial, antimalarial, and antitumor activity. Recently, increasing experimental studies demonstrated that many Artemisia extracts offer a great antiepileptic potential, which was attributed to their bioactive components via various mechanisms of action. However, detailed literature on the antiepileptic properties of the genus Artemisia and its mechanism of action is segregated. In this review, we tried to gather the detailed neuroprotective and antiepileptic properties of the genus Artemisia and its possible underlying mechanisms. In this respect, 63 articles were identified in the PubMed and Google scholars databases, from which 18 studies were examined based on the pharmacological use of the genus Artemisia species in epilepsy. The genus Artemisia extracts have been reported to possess antioxidant, anti-inflammatory, neurotransmitter-modulating, anti-apoptotic, anticonvulsant, and pro-cognitive properties by modulating oxidative stress caused by mitochondrial ROS production and an imbalance of antioxidant enzymes, by protecting mitochondrial membrane potential required for ATP production, by upregulating GABA-A receptor and nACh receptor activities, and by interfering with various anti-inflammatory and anti-apoptotic signaling pathways, such as mitochondrial apoptosis pathway, ERK/CREB/Bcl-2 pathway and Nrf2 pathway. This review provides detailed information about some species of the genus Artemisia as potential antiepileptic agents. Hence, we recommend further investigations on the purification and identification of the most biological effective compounds of Artemisia and the mechanisms of their action to cure epilepsy and other neurological diseases.

Shisa7-Dependent Regulation of GABAA Receptor Single-Channel Gating Kinetics.

GABAA receptors (GABAARs) mediate the majority of fast inhibitory transmission throughout the brain. Although it is widely known that pore-forming subunits critically determine receptor function, it is unclear whether their single-channel properties are modulated by GABAAR-associated transmembrane proteins. We previously identified Shisa7 as a GABAAR auxiliary subunit that modulates the trafficking, pharmacology, and deactivation properties of these receptors. However, whether Shisa7 also regulates GABAAR single-channel properties has yet to be determined. Here, we performed single-channel recordings of α2β3γ2L GABAARs cotransfected with Shisa7 in HEK293T cells and found that while Shisa7 does not change channel slope conductance, it reduced the frequency of receptor openings. Importantly, Shisa7 modulates GABAAR gating by decreasing the duration and open probability within bursts. Through kinetic analysis of individual dwell time components, activation modeling, and macroscopic simulations, we demonstrate that Shisa7 accelerates GABAAR deactivation by governing the time spent between close and open states during gating. Together, our data provide a mechanistic basis for how Shisa7 controls GABAAR gating and reveal for the first time that GABAAR single-channel properties can be modulated by an auxiliary subunit. These findings shed light on processes that shape the temporal dynamics of GABAergic transmission.SIGNIFICANCE STATEMENT Although GABAA receptor (GABAAR) single-channel properties are largely determined by pore-forming subunits, it remains unknown whether they are also controlled by GABAAR-associated transmembrane proteins. Here, we show that Shisa7, a recently identified GABAAR auxiliary subunit, modulates GABAAR activation by altering single-channel burst kinetics. These results reveal that Shisa7 primarily decreases the duration and open probability of receptor burst activity during gating, leading to accelerated GABAAR deactivation. These experiments are the first to assess the gating properties of GABAARs in the presence of an auxiliary subunit and provides a kinetic basis for how Shisa7 modifies temporal attributes of GABAergic transmission at the single-channel level.

GABAA receptors as targets for anaesthetics and analgesics and promising candidates to help treat coronavirus infections: A mini-review.

GABA is a major inhibitory neurotransmitter that regulates the balance between excitatory and inhibitory circuits in the human nervous system. The GABA receptors are divided into three main subtypes, GABAA, GABAB, and GABAC (also termed GABAA rho) receptors. GABAA receptors are pentameric ligand‐gated ion channels widely expressed throughout the central and peripheral nervous system. The activation of GABAA receptors results in opening of an anion‐selective channel that mainly gates chloride ions and allows them to flow into the neuron, causing hyperpolarization of the cell membrane that dampens neural excitability. This makes GABAA receptors critical anaesthetic and analgesic targets for existing as well as for the development of novel drugs. In this review, we first summarize the biochemical properties of GABAA receptors and the clinical anaesthetics and analgesics targeting the receptors. In a forward‐looking section, we summarize the emerging role of GABAergic signalling in treatment of COVID‐19 related infections. Finally, we discuss the opportunities arising from targeting specific and unique subunit interfaces for the development of novel anaesthetics and analgesics leading to more efficient therapies.